RESUMO
Diterpenes are an important class of plant metabolites that can be used in the search for new antibacterial agents. ent-Copalic acid (CA), the major diterpene in Copaifera species exudates, displays several pharmacological properties. This study evaluates the CA antibacterial potential against the anaerobic bacteria Peptostreptococcus anaerobius and Actinomyces naeslundii. Antimicrobial assays included time-kill and biofilm inhibition and eradication assays. Time-kill assays conducted for CA concentrations between 6.25 and 12.5⯵g/mL evidenced bactericidal activity within 72â¯h. CA combined with chlorhexidine dihydrochloride (CHD) exhibited bactericidal action against P. anaerobius within 6â¯h of incubation. As for A. naeslundii, the same combination reduced the number of microorganisms by over 3 log10â¯at 24â¯h and exerted a bactericidal effect at 48â¯h of incubation. CA at 500 and 2000⯵g/mL inhibited P. anaerobius and A. naeslundii biofilm formation by at least 50%, respectively. CA at 62.5 and 1.000⯵g/mL eradicated 99.9% of pre-formed P. anaerobius and A. naeslundii biofilms, respectively. These results indicated that CA presents in vitro antibacterial activity and is a potential biofilm inhibitory agent. This diterpene may play an important role in the search for novel sources of agents that can act against anaerobic bacteria.
Assuntos
Actinomyces/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Diterpenos/farmacologia , Peptostreptococcus/efeitos dos fármacos , Extratos Vegetais/farmacologia , Actinomyces/fisiologia , Fabaceae/química , Testes de Sensibilidade Microbiana , Peptostreptococcus/fisiologiaRESUMO
In view of the biological activities and growing therapeutic interest in oleoresin obtained from Copaifera multijuga, this study aimed to determine the genotoxic and antigenotoxic potential of this oleoresin (CMO) and its chemical marker, diterpene (-)-copalic acid (CA). The micronucleus (MN) assay in V79 cell cultures and the Ames test were used for in vitro analyses, as well as MN and comet assays in Swiss mice for in vivo analyses. The in vitro genotoxicity/mutagenicity results showed that either CMO (30, 60, or 120 µg/ml-MN assay; 0.39-3.12 mg/plate-Ames test) or CA (2.42; 4.84, or 9.7 µg/ml-MN assay; 0.39-3.12 mg/plate-Ames test) did not induce a significant effect on the frequency of MN and number of revertants, demonstrating an absence of genotoxic and mutagenic activities, respectively, in vitro. In contrast, these natural products significantly reduced the frequency of MN induced by methyl methanesulfonate (MMS), and exerted a marked inhibitory effect against indirect-acting mutagens in the Ames test. In the in vivo test system, animals treated with CMO (6.25 mg/kg b.w.) exhibited a significant decrease in rate of MN occurrence compared to those treated only with MMS. An antigenotoxic effect of CA was noted in the MN test (1 and 2 mg/kg b.w.) and the comet assay (0.5 mg/kg b.w.). Data suggest that the chemical marker of the genus Copaifera, CA, may partially be responsible for the observed chemopreventive effect attributed to CMO exposure. ABBREVIATIONS: 2-AA, 2-anthramine; 2-AF, 2-aminofluorene; AFB1, aflatoxin B1; B[a]P, benzo[a]pyrene; BOD, biological oxygen demand; BPDE, benzo[a]pyrene-7,8-diol-9,10-epoxide; CA, (-)-copalic acid; CMO, oleoresin of Copaifera multijuga, DMEM, Dulbecco`s Modified Eagles`s Medium; DMSO, dimethylsulfoxide; EMBRAPA, Brazilian agricultural research corporation; GC-MS, gas chromatography-mass spectrometry; HAM-F10, nutrient mixture F-10 Ham; HPLC, high performance liquid chromatography; LC-MS, liquid chromatography-mass spectrometry; MI, mutagenic index; MMC, mitomycin C; MMS, methyl methanesulfonate; MN, micronucleus; MNPCE, micronucleated polychromatic erythrocyte; NCE, normochromatic erythrocyte; NDI, nuclear division index; NMR, nuclear magnetic resonance; NPD, 4-nitro-o-phenylenediamine; PBS, phosphate-buffered saline; PCE, polychromatic erythrocyte; SA, sodium azide; V79, Chinese hamster lung fibroblast.
Assuntos
Antimutagênicos/farmacologia , Diterpenos/farmacologia , Fabaceae/química , Extratos Vegetais/farmacologia , Animais , Ensaio Cometa , Cricetulus , Fibroblastos/efeitos dos fármacos , Pulmão , Masculino , Camundongos , Testes para Micronúcleos , Testes de MutagenicidadeRESUMO
Copaiba oleoresins are used in alternative medicine as anti-inflammatory, antitumoral, and antimicrobial treatments. (-)-Copalic acid (CA) is the major diterpene found in exudates from Copaifera species. We have examined the genotoxicity and the chemopreventive potential of Copaifera multijuga oleoresin (CM) and CA. Genotoxicity assessment was examined with the peripheral blood micronucleus test and the comet assay (male Swiss mouse hepatocytes). In the chemoprevention study, we evaluated the effects of CM and CA on the formation of 1,2-dimethylhydrazine (DMH)-induced aberrant crypt foci (ACF) in male Wistar rat colon. Neither agent caused a significant increase in micronucleus frequency relative to controls, but the highest CM dose tested (400mg/kg b.w.) caused DNA damage in the comet assay. Both agents significantly reduced the frequency of DMH-induced ACF. Both CM and CA suppressed ACF formation and may have a protective effect against colon carcinogenesis.
Assuntos
Anticarcinógenos/farmacologia , Dano ao DNA , Diterpenos/farmacologia , Fabaceae/química , Micronúcleos com Defeito Cromossômico/induzido quimicamente , Extratos Vegetais/farmacologia , Focos de Criptas Aberrantes/prevenção & controle , Animais , Anticarcinógenos/isolamento & purificação , Anticarcinógenos/toxicidade , Neoplasias do Colo/prevenção & controle , Ensaio Cometa , Diterpenos/isolamento & purificação , Diterpenos/toxicidade , Relação Dose-Resposta a Droga , Hepatócitos/efeitos dos fármacos , Hepatócitos/patologia , Masculino , Camundongos , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/toxicidade , Ratos WistarRESUMO
We report herein the synthesis of six diterpene derivatives, three of which are new, generated through known organic chemistry reactions that allowed structural modification of the existing natural products kaurenoic acid (1) and copalic acid (2). The new compounds were fully characterized using high resolution mass spectrometry, infrared spectroscopy, ¹H- and (13)C-NMR experiments. We also report the evaluation of the anti-tuberculosis potential for all compounds, which showed some promising results for Micobacterium tuberculosis inhibition. Moreover, the toxicity for each of the most active compounds was also assessed.